Clinical isolates of staphylococci (Staphylococcus aureus and S. epidermidis) which cause serious infections due to their intrinsic resistance to beta-lactamase-stable beta-lactam antibiotics (e.g., methicillin) carry the mecA gene. Song et al., FEBS Lett. 221:167-171 (1987). This gene encodes a putative cell wall biosynthetic enzyme referred to as penicillin binding protein 2a (PBP2a). PBP2a, which binds beta-lactams only at concentrations well above therapeutic efficacy, apparently can functionally substitute for all the staphylococcal PBPs and permit growth when the host organism is threatened by beta-lactams. Hartman and Tomasz, J. Bacteriol. 158:513-516 (1984). Wu et al., Antimicrob. Agents Chemother. 36:533-539 (1992) and Ryffel et al., Gene 94:137-138 (1990).
The mecA gene is not a normal part of the staphylococcal genome. The organism which donated mecA to the staphylococci remains unidentified. Despite the uniform presence of mecA in methicillin-resistant clinical isolates, these isolates vary considerably in their degree of resistance to methicillin. This variation in phenotypic expression within a population has been referred to as heterogenous expression. Matthews and Stewart, FEMS Microbiol. Lett. 22:161-166 (1984). Typically, most cells exhibit low-level resistance to methicillin and only a minority of the population express high-level resistance, perhaps only one in 10.sup.8 cells. Tomasz et al., Antimicrob. Agents Chemother. 35:124-129 (1991). Although expression of methicillin resistance is dependent upon the presence of PBP2a, it appears to be somewhat independent of the amount of PBP2a, suggesting important roles for other factors. Chambers and Hackbarth, Antimicrob. Agents Chemother. 31:1982-1988 (1987) and Murakami and Tomasz, J. Bacteriol. 171:874-879 (1989).
Tn551 insertional mutagenesis of methicillin-resistant S. aureus revealed numerous sites which influence the level of methicillin resistance but are non linked to mecA and do not perturb the expression of PBP2a. Berger-B achi et al., Antimicrob. Agents Chemother. 36:1367-1373 (1992); Kornblum et al., Eur. J. Clin. Microbiol. 5:714-718 (1986); Berger-B achi et al., Mol. Gen. Genet. 219:263-269 (1989) and Maidhof et al., J. Bacteriol. 173:3507-3513 (1991). Those factors described thus far generally depress the MIC of beta-lactam resistant strains. Some of the genetic loci which demonstrate such an effect on methicillin resistance were designated factors essential for methicillin resistance (fem). Berger-B achi et al., Mol. Gen. Genet. 219:263-269 (1989). In contrast to mecA, the genes which encode influential factors are probably present in both resistant and susceptible strains of S. aureus and S. epidermidis. Information obtained from gene disruption studies of femA and femB in S. aureus indicated that in addition to enhanced sensitivity to methicillin, homogeneously methicillin-resistant S. aureus strains carrying such gene disruptions have a reduced glycine content in the peptidoglycan component of their cell walls (Maidhof et al., J. Bacteriol. 173:3507-3513 (1991)) and exhibit reduced rates of cell wall turnover and autolysis (de Jonge et al., J. Bacteriol. 173:1105-1110 (1991)).
Genetic factors, other than mecA, that influence the expression of methicillin resistance in S. epidermidis have, until now, not been described at the molecular level. The present invention provides DNA sequences encoding the FemA protein of Staphylococcus epidermidis, the FemA protein itself, and vectors and microorganisms comprising the femA gene of S. epidermidis.